A droplet actuator typically includes one or more substrates configured to form a surface or gap for conducting droplet operations. The one or more substrates establish a droplet operations surface or gap for conducting droplet operations and may also include electrodes arrange to conduct the droplet operations. The droplet operations substrate or the gap between the substrates may be coated or filled with a filler fluid that is immiscible with the liquid that forms the droplets.
Droplet actuators are used in a variety of applications, including molecular diagnostic assays such as enzymatic assays and immunoassays. In one application, enzymatic assays and immunoassays are used as part of a routine testing process to test newborn infants for various genetic disorders. For example, enzymatic assays may be used to test for various lysosomal storage diseases (LSD). Many genetic disorders targeted in newborn testing programs are associated with deficiencies in glycoprotein molecules (e.g., lysosomal enzymes) that may represent only a small fraction of total macromolecules in a complex biological sample, such as blood. Samples for newborn testing are typically collected by pricking the heel of the newborn to obtain a small quantity of blood (typically, two to three drops) to fill a few circles on a filter paper card. Specimens are punched from the DBS sample (i.e., one punch for each test to be performed using current technologies) and manipulated according to a specific assay protocol. Because the blood samples are spotted onto a solid medium and dried, they must be reconstituted before analysis, a step that requires dilution of the sample into a suitable liquid medium. There is a need for improved methods for increasing sensitivity of detection of target glycoprotein molecules in NBS assays.